The protozoan parasite Trypanosoma cruzi (T. cruzi) is the causative agent of Chagas disease, and is a prominent human health problem in Central and South America. The burden of Chagas disease is estimated to be five to ten times greater than that of malaria. Chagas disease results from chronic infection with T. cruzi, and affects up to 10 million people worldwide: of those 50,000 die each year. Global migration patterns have contributed to the spread of T. cruzi infection in the United States, Europe, and Japan, and this is likely to continue. Despite its enormous impact on public health, control of T. cruzi infection s hampered by the absence of protective or therapeutic vaccines. Even though the role of the immune T cell response in the control of Chagas disease has been clearly demonstrated, the identification of the T. cruzi antigens that elicit these immune responses in patients is still ver limited. Furthermore, in the last few years several studies have focused on the identification of immunogenic T. cruzi antigens that could be used as candidates for the development of a Chagas vaccine. While varying levels of protective immunity have been accomplished with a number of antigens, it is clear that new antigens could contribute to develop new vaccine formulations. In addition, it is important to note that none of the antigens that have been evaluated as vaccines have been derived from the examination of the specificity of T cells from T. cruzi infected individuals. [We propose a proof-of-concept study focused on the generation of memory CD4+ T cell libraries and clones from patients with Chagas disease and the identification of the antigen specificities using positional scanning peptide libraries. T cell cloes from indeterminate/asymptomatic subjects will be used for the identification of immunogenic T. cruzi antigens since these patients have remained, for more than 30 years, without detectable clinical manifestations of disease, and therefore are likely to recognize parasite antigens involved in protection. The recognition and immunoprevalence of the identified peptides will be evaluated with T cell libraries from indeterminate/asymptomatic subjects as well as chronic cardiac patients and non-infected individuals to determine the differences in their recognition and to further understand their relevance in protection and the progression of the disease]. The results of the proposed studies will advance the identification of the antigens recognized by infected individuals and contribute in the design of new prophylactic and/or therapeutic Chagas vaccines.